Project summary Sudden cardiac death (SCD), caused by severe arrhythmias such as ventricular tachycardia (VT) and ventricular fibrillation (VF), claims the lives of 300,000-400,000 Americans annually. SCD often occurs during acute myocardial ischemia or infarction. The mechanism(s) and prevention of ischemia-induced VT/VF and SCD remain to be important topics in both basic research and clinics. It has been shown that the hearts of hibernating animals are resistant to cardiac arrhythmia (including VF) induced by many interventions such as hypothermia, Ca-overload, and also ischemia, etc. Supporting these observations, our preliminary results demonstrated seasonal variation in the antiarrhythmic properties in hibernating mammals, specifically, the woodchucks (Marmota monax) in winter are more resistant to ischemia-induced VF and SCD than woodchucks in summer. It seems likely that hibernating mammals (such as woodchucks) undergo intrinsic adaptive processes to prepare for winter hibernation, which would protect them against cardiac injury and arrhythmias (VF and SCD). Although adaptive alternations in membrane ion channels, Ca handling, connexin expression, and signaling molecules have been reported in hibernating animals, the direct mechanistic link between adaptation of cellular and ionic properties and antiarrhythmic protection in hibernators is still unclear. Reactive oxygen species (ROS) are released acutely in large amounts during ischemia or ischemia/reperfusion injury. Pathological concentrations of H2O2 causes early afterdepolarization (EAD), delayed afterdepolarization (DAD), and triggered activity (TA), and has been suggested as a potential proarrhythmic factor in rats, guinea pigs, and rabbits. Our recent studies have revealed that H2O2 induces EAD, DAD and TA via activation of the Ca/Calmodulin-dependent kinase II (CaMKII) pathway in rabbit ventricular myocytes. Our preliminary studies also showed up-regulation of antioxidant enzymes, e.g., MnSOD and lower levels of CaMKII activity (oxidized form) in woodchuck hearts in winter than in summer. Taken together, these findings led us to hypothesize that woodchucks undergo natural adaptations in winter by reducing oxidative stress (ROS levels) and thus ROS- induced (oxidized) CaMKII activity levels, which benefits the woodchucks and confer resistance against ischemia-induced VF and SCD in winter. The major goal of this proposal is to investigate the molecular and cellular mechanisms involved in the resistance against arrhythmia in woodchucks in winter compared to in summer (and to non-hibernators) by determining the signaling molecules and examining their susceptibility to H2O2 induced EADs, DADs and TAs. We will assess Hypothesis 1 (in intact heart and at the tissue level): Woodchuck hearts exhibit greater resistance to ischemia-induced VF and SCD in winter. Lower oxidative stress (ROS) levels and ROS-oxidized CaMKII activity levels account for the antiarrhythmic mechanism in woodchucks in winter; and Hypothesis 2 (in isolated single myocytes): Isolated myocytes from winter woodchucks exhibits less vulnerability to ROS-induced EAD and TA in a CaMKII dependent manner. The hibernating mammals represent a novel model, which may provide insights into understanding and prevention of ischemia-related arrhythmias and SCD.